The studies in the current project are directed at solving the problems that must be surmounted to refine the recently demonstrated unique clinical potentials of intradermal (I.D.) gene vaccination into a practical and efficacious form of immunotherapy. In Aim 1, the various regulatory, structural and immune modulatory components within plasmid-based vectors will be systematically evaluated to identify the variations with the optimal properties for gene immunization. In part, these studies will consider different means for increasing and targeting the expression of cDNA protein to different cellular and extracellular compartments. Specific non-encoding DNA elements will also be evaluated for their utility to augment desirable immune responses and evaluate strategies for improving the safety of these vectors for human vaccination. In Aim 2, we will seek to solve the unique problems associated with applying DNA vaccination for the immunotherapy of environmental allergies. cDNA constructs will be designed and tested to evaluate for their ability to induce clinically desirable responses to the house dust mite, Dermatophagoides pteronyssinus, (Der p) in the mouse. These studies will evaluate whether immunization with a gene encoding a truncated Der p gene can induce a Th1 switch that ultimately "spreads" to encompass the entire protein and to other structurally distinct proteins. In Aim 3, phage-display technology currently in use in our laboratory will be used to efficiently and rapidly isolate specific cDNA that encode for allergen proteins based on recognition by allergic IgE from atopic individuals. The insights and materials obtained from these studies will contribute approaches that will also assist the advancement of other projects in the current program.